Ex vivo lung-organoid model for aberrant basaloid cell induction and activation

Lung epithelial cell injury contributes to various lung diseases, impacting lung function and overall human health. Bleomycin (BLM)-induced murine models of pulmonary fibrosis (PF) are commonly utilized to study lung epithelial cell injury. However, the complex microenvironments in inflamed lungs hinder clear causal relationships between cell-to-cell, molecules interaction. To address this limitation, we aimed to establish an ex vivo organoid model of BLM-induced lung injury recapitulating the cellular and molecular responses of BLM-induced lung epithelial injury in vivo. Overall design: Lung organoids were generated by co-culturing primary murine CD326 positive lung epithelial cells and CD140a positive lung fibroblasts. To clarify transcriptomic changes of BLM-injured lung organoids at single-cell level, we stimulated lung organoids with bleomycin (0.3 ug/ml) in a culture media from day 8 to day 10 post co-cultured. After that, bleomycin was removed from the culture media and the lung organoids were further cultured until day 14 post co-cultured. scRNA-seq analysis was performed at day 14 post co-cultured.